人肾间质成纤维细胞体外增殖及胶原的合成

In vitro proliferation and collagen synthesis of human renal interstitial fibroblasts

背景:肾间质成纤维细胞体外研究时间点的选取多在72h以内,但对于体外长时间动态观察培养过程中成纤维细胞增殖及胶原分泌规律少见报道。目的:观察人肾间质纤维化来源的成纤维细胞体外增殖及胶原蛋白合成的规律。方法:采用组织块培养法获取、纯化并鉴定肾间质纤维化来源的人肾间质成纤维细胞,选取第3代对数生长期细胞用于实验。MTT比色法及Masson三色染色结合图像分析法分别检测体外培养第1,3,5,7,9天细胞的吸光度值和成纤维细胞胶原蛋白染色阳性细胞的平均吸光度值。结果与结论:肾间质成纤维细胞体外生长曲线呈"S"形,体外培养第3～5天为细胞倍增时间,随培养时间延长,胶原蛋白由线状分布转变成片状分布,其中以第3～5天增长最快(P<0.05)。结果证实,肾间质成纤维细胞在体外培养第3～5天增殖能力和胶原合成活性最强。

BACKGROUND：The time points for in vitro studies about renal interstitial fibroblasts are mainly selected within 72 hours.However,there are rare reported about proliferation and collagen synthesis of renal interstitial fibroblasts cultured in vitro for a long term.OBJECTIVE：To investigate the pattern of proliferation and collagen synthesis of human renal interstitial fibroblasts in vitro.METHODS：human renal interstitial fibroblasts were obtained,purified and identified.The third passage cells were used in experiment.Cell proliferation was determined by MTT method and collagen synthesis was observed using Masson trichrome staining accompanied with image analysis at the 1^st,3^rd,5^th,7^th and 9^th days after culture in vitro.RESULTS AND CONCLUSION：The growth curve of renal interstitial fibroblasts was shown as ＂S＂ and doubling time was from the 3^rd to 5^th day in vitro;with the culturing time extending,the distribution of collagen protein changed from line into lamellar and the quantity of collagen protein increased obviously from the 3^rd to 5^th day （P〈0.05）.It is indicated that the biological activity of renal interstitial fibroblasts is the most productive from the 3^rd to 5^th day,which might be an important target during the course of pharmacological experiment in vitro.